Reduction of Escherichia coli O157:H7 in cattle drinking-water by trans-cinnamaldehyde

Cattle serve as a major reservoir of E. coli O157:H7 and excrete the pathogen in feces. Environmental persistence of E. coli O157:H7 plays a vital role in its epidemiology on farms, and cattle water troughs are a demonstrated long-term reservoir of E. coli O157:H7 for animals. The objective of this study was to investigate the potential of low concentrations of trans-cinnamaldehyde for killing E. coli O157:H7 in cattle drinking-water. A five-strain mixture of E. coli O157:H7 was inoculated (at approximately 8.0 log colony-forming units [CFU]/mL) into 100 mL samples of well water containing 0, 0.03, 0.05, 0.07, or 0.1% trans-cinnamaldehyde. Additionally, water samples containing (1% w/v) bovine feces or feed were also included. The samples were incubated at 21 degrees , 8 degrees , or 4 degrees C for 7 days and tested for viable E. coli O157:H7 on days 0, 1, 3, 5, and 7. Triplicate samples of each treatment and control were included and the study was replicated twice. All concentrations of trans-cinnamaldehyde were effective in killing E. coli O157:H7 in water, but the magnitude of killing significantly increased with increase in trans-cinnamaldehyde concentration and storage temperature (p < 0.05). The presence of feed or feces in water decreased the antibacterial effect of trans-cinnamaldehyde on E. coli O157:H7 (p < 0.05). This study indicated that trans-cinnamaldehyde is effective in killing E. coli O157:H7 in cattle drinking-water, but detailed palatability studies on cattle intake of water containing the antimicrobial are needed.     

Evaluation of hha and hha sepB mutant strains of Escherichia coli O157:H7 as bacterins for reducing E. coli O157:H7 shedding in cattle

Escherichia coli O157:H7 colonizes cattle intestines by using the locus of enterocyte effacement (LEE)-encoded proteins. The induction of systemic immune response against LEE-encoded proteins, therefore, will prove effective in reducing E. coli O157:H7 colonization in cattle. The previous studies have demonstrated that a hha (encodes for a hemolysin expression modulating protein) deletion enhances expression of LEE-encoded proteins and a sepB (encodes an ATPase required for the secretion of LEE-encoded proteins) deletion results in intracellular accumulation of LEE proteins. In this study, we demonstrate the efficacy of the hha and hha sepB deletion mutants as bacterins for reducing fecal shedding of E. coli O157:H7 in experimentally inoculated weaned calves. The weaned calves were injected intramuscularly with the bacterins containing 10⁹ heat-killed cells of the hha⁺ wild-type or hha or hha sepB isogenic mutants, and boosted with the same doses 2- and 4-weeks later. The evaluation of the immune response two weeks after the last booster immunization revealed that the calves vaccinated with the hha mutant bacterin had higher antibody titers against LEE proteins compared to the titers for these antibodies in the calves vaccinated with the hha sepB mutant or hha⁺ wild-type bacterins. Following oral inoculations with 10¹⁰ CFU of the wild-type E. coli O157:H7, the greater numbers of calves in the group vaccinated with the hha or hha sepB mutant bacterins stopped shedding the inoculum strain within a few days after the inoculations compared to the group of calves vaccinated with the hha⁺ wild-type bacterin or PBS sham vaccine. Thus, the use of bacterins prepared from the hha and hha sepB mutants for reducing colonization of E. coli O157:H7 in cattle could represent a potentially important pre-harvest strategy to enhance post-harvest safety of bovine food products, water and produce.     

Seasonal shedding of Escherichia coli O157:H7 in ruminants: a new hypothesis

Seasonal shedding of Escherichia coli O157:H7 in ruminants is well documented; however, viable explanations for this phenomenon are lacking. The fecal prevalence of E. coli O157:H7 in ruminants is the highest in the summer months, decreasing to low or undetectable levels in the winter. To determine if day length is correlated to seasonal shedding, nine reports of E. coli O157:H7 prevalence in cattle were selected for similarity of sampling technique, culture methodology, cattle type, and ability to determine approximate sampling month and location, representing three regions in North America (southern Canada, midwestern United States, and the southern United States/Mexico). Correlation and regression analysis revealed a positive correlation existed between day length (r = 0.67; R2 = 0.45; P = 0.0009) and, to a lesser extent, ambient temperature (r = 0.43; R2 = 0.19; P = 0.05) and E. coli O157:H7 prevalence. Based on these results, a study was conducted in a commercial feedlot to examine the effect of artificial lighting on E. coli O157 prevalence. Four pens of cattle received approximately 5 hours of artificial lighting daily (in addition to the natural light) for 60 days and four pens served as controls. Freshly voided fecal samples (representing approximately 30% of the steers/pen) were collected from the pen floor and cultured for E. coli O157 after 25 and 53 days of artificial lighting and 28 and 43 days following cessation of the lighting treatment. Fecal prevalence of E. coli O157 remained constant in the lighted pens whereas prevalence was lower (P < 0.01) in the control treatment after 53 days of lighting. Forty-three days following cessation of the lighting treatment, prevalence decreased in the lighted treatment group to levels comparable (P > 0.20) to control levels. Understanding the mechanisms underlying the seasonality of this important foodborne pathogen will enable the implementation of effective control strategies when it is most prevalent.     

Swimming-associated outbreak of Escherichia coli O157:H7

In 1997 the first outbreak of Escherichia coli O157:H7 infections involving 14 cases occurred in Finland. A case was defined as a resident of Alavus with an episode of diarrhoea between 5 and 17 July 1997, and from whom E. coli O157:H7 was isolated from stool. The investigation included case searching and a population-based case control study. Five primary and eight symptomatic secondary cases of E. coli O157:H7 illness were detected. In the 10 days before the outbreak, all 5 primary patients (aged 3-8 years), but only 6 of 32 population controls from the same age range (Fisher's test, P < 0.001) and 4 of 10 sibling controls (P < 0.05) had visited (but had not necessarily bathed in) a shallow beach popular among young children. Four out of 5 primary cases had remained within 5 m of the beach while swimming and had swallowed lake water compared to 1 of 5 population controls. These analytical epidemiologic findings incriminated fresh lake water as the vehicle of E. coli O157:H7 transmission.     

Chlorine inactivation of Escherichia coli O157:H7.

We analyzed isolates of Escherichia coli O157:H7 (which has recently caused waterborne outbreaks) and wild-type E. coli to determine their sensitivity to chlorination. Both pathogenic and nonpathogenic strains were significantly reduced within 1 minute of exposure to free chlorine. Results indicate that chlorine levels typically maintained in water systems are sufficient to inactivate these organisms.     

从农村耕牛粪便中检出产志贺毒素大肠埃希菌O157:H7

目的：调查东阳市家畜、家禽中O157：H7的携带状况,并进一步了解分离到的1株STEC O157：H7的分子生物学特性。方法：随机采集全市各乡镇的家畜、家禽粪便及其生鲜肉样品,进行大肠埃希菌O157：H7的检测,应用O157和H7特异性抗血清凝集试验进行菌株血清型的鉴定;选用ATB全自动微生物鉴定仪和VITEK 2-COMPACT进行常规生化及药敏试验;使用PCR检测O、H抗原编码基因和SLTl、SLT2、hly、eaeA 4种毒力基因。结果：在109份样品中,分离出1株经细菌生化鉴定为大肠埃希菌O157,血清型为O157：H7的菌株;使用PCR检测到O157：H7的O、H抗原编码基因和毒力基因SLT2、hly、eaeA,但未检测到毒力基因SLTl。结论：从东阳市的一头农村耕牛粪便中分离到1株STEC O157：H7,提示疾控中心要加强主动监测,及时发现可能发生的疫情,通过有效干预,防范肠出血性大肠杆菌O157：H7所致食源性疾病的发生。     

2004-2007年云南省肠出血性大肠埃希菌0157:H7监测分析

肠出血性大肠埃希菌O157:H7(O157:H7)是近年来新发现的危害严重的肠道致病菌.自1982年美国首次暴发O157:H7,感染性腹泻以来,世界各地陆续有疫情出现[1,2].     

Extended phage-typing scheme for Escherichia coli O157:H7

In Canada, the number of human isolates of verotoxigenic (VT + ve) Escherichia coli O157:H7 from diarrhoeal cases and haemolytic uraemic syndrome and haemorrhagic colitis has increased from 25 in 1982 to 2384 in 1989. A total of 3273 VT + ve E. coli O157:H7 strains (3255 strains isolated in Canada and 18 isolates from other countries) were phage typed. The phage typing scheme has been extended from 14 to 62 phage types. Of these, five types occurred exclusively in other countries (type 47 in Japan; and types 49, 50, 51 and 52 in the U.K.). Thirty-five different phage types were identified in Canada; only nine of these (1, 2, 4, 8, 14, 21, 23, 31 and 32), each accounted for more than 1% of the cases from human sources. The same nine types were the only ones observed among the isolates from non-human sources (meat and slaughter houses) suggesting a food-borne transmission in most of the human cases. Phage types 1 (30.5%); 4 (21%); 8 (13.5%); 31 (8.9%) and 14 (8%) were encountered in varying frequencies in most of the provinces; infrequently occurring phage types also showed regional variation. Thirteen different phage types were identified among 151 outbreaks representing 556 isolates of E. coli O157:H7. More than one phage type were encountered in 12 outbreaks whereas in 141 outbreaks, all strains in each, had the same phage type.     

The shift of genetic subtypes of Escherichia coli O157:H7 isolates from cattle.

A total of 46 Escherichia coli O157:H7 isolates were obtained from sequential faecal samples from seven cattle collected over periods of 2 months. Nine closely related genetic subtypes, determined by pulsed-field gel electrophoresis types using three kinds of restriction endonuclease were observed among the isolates. Distinguishable, but closely related genetic subtypes can be isolated from one farm, or from one cow, should be considered when undertaking an epidemiological survey.     

环介导等温扩增法应用于肠出血性大肠埃希菌O157:H7的检测

的仪器,非常适合基层检验部门及小型实验室与流行病学人员现场监测使用,具有较为广泛地应用前景.     

A two-dose regimen of a vaccine against Escherichia coli O157:H7 type III secreted proteins reduced environmental transmission of the agent in a large-scale commercial beef feedlot clinical trial

A clinical vaccine trial of commercially fed cattle tested the effect of a two-dose regimen of a vaccine product against type III secreted proteins of enterohemorrhagic Escherichia coli O157:H7 on the probability of detecting the organism on environmental sampling devices. Within commercial feedlots, pens of vaccinated and unvaccinated cattle were matched by reprocessing schedule and time of sampling. Vaccine was administered to all cattle within treated pens at arrival processing and again at re-implant processing. Pens of cattle were sampled 1 week after the second dose of vaccine and every 3 weeks for four test periods. Pair-matched pens of cattle were sampled concurrently. Test samples were seven ropes per pen hung overnight from the feed-bunk neck-rail (ROPES). Recovery of E. coli O157:H7 from at least one rope classified pens ROPES-positive. E. coli O157:H7 isolates were identified by standard biochemical methods and multiplex polymerase chain reaction. The probability for pens of cattle to test ROPES-positive was modeled using multilevel logistic regression with variance adjustment for clustering by matched pens and repeated measures. We studied 140 pens of cattle representing 20,556 cattle in 19 feedlots February through October 2004. Vaccinated pens of cattle were less likely to test ROPES-positive (OR = 0.59, p = 0.004). Because ROPES testing identifies organisms in the mouth of cattle, and the outcome is both associated with presence of the organism in the pen environment and correlated with the prevalence of fecal shedding, we conclude the two-dose vaccine regimen reduces the probability for environmental transmission of E. coli O157:H7 within commercial cattle feeding systems.     

Hemolytic uremic syndrome risk and Escherichia coli O157:H7

We reviewed medical records of 238 hospitalized patients with Escherichia coli O157:H7 diarrhea to identify risk factors for progression to diarrhea-associated hemolytic uremic syndrome (HUS). Data indicated that young age, long duration of diarrhea, elevated leukocyte count, and proteinuria were associated with HUS.     

Cross reactivity of enterohemorrhagic Escherichia coli O157:H7-specific sera with non-O157 serotypes

Escherichia coli O157:H7 is an important food- and water-borne pathogen of humans, causing Hemorrhagic Colitis and Haemolytic Uremic Syndrome. Colonization of both cattle and human hosts is mediated through the action of effector molecules secreted via a Type III secretion system, a mechanism shared by other enterohemorrhagic E. coli (EHEC). We recently reported that vaccination of cattle with Type III-secreted proteins (TTSPs) resulted in decreased shedding of the organism following both experimental infection as well as under conditions of natural exposure. In order to extend this to non-O157 EHEC serotypes, we examined the serological cross reactivity of TTSPs of serotypes O26:H11, O103:H2, O111:NM and O157:H7. Western blotting experiments with polyclonal antisera directed against serotype O157:H7 TTSPs suggested that there was significant cross reactivity, although there was limited cross reactivity when two Tir- and EspA-specific monoclonal antibodies were used. Groups of cattle were then vaccinated with TTSPs produced from each of the above serotypes and the magnitude and specificity of the responses were measured. All animals responded well with antibodies to TTSPs of the homologous serotype. However, limited cross reactivity was observed against the others. No cross reactivity was observed against Tir and EspA of serotype O157:H7. These results suggest that vaccination of cattle with TTSPs as a means of reducing the risk of EHEC transmission to humans will induce protection that is serotype specific.     

Persistence of Escherichia coli O157:H7 in dairy cattle and the dairy farm environment.

The persistence of Escherichia coli O157:H7 in cattle and the farm environment was investigated on eight Ontario dairy farms positive for E. coli O157:H7 in a longitudinal study commenced one year previously. Faecal samples from cows, calves, humans, cats, rodents, wild birds, a composite fly sample and numerous composite and individual environmental samples were cultured and tested for verotoxin-producing E. coli (VTEC). VTEC isolates were serotyped and E. coli O157:H7 isolates were phage typed. E. coli O157:H7 phage type 34 was isolated from one calf on each of two farms. The same phage type had been isolated on one of these farms 12 months earlier. Most E. coli O157:H7-positive animals and farms became culture-negative within 2 and 3 months, respectively. E. coli O157:H7 was not isolated from any environmental samples, although evidence of VTEC was found in composite samples from calf feeders (19.1%), calf barn surfaces (18%), cow feeders (14.9%), flies (12.5%), cow barn surfaces (11.3%), and individual milk filters (12.5%). VTEC belonging to 21 non-O157 serotypes were isolated from 24 cows (8.2%), 21 calves (18.3%), 2 cow feeder samples (3.0%), and 1 calf feeder sample (4.8%). Shedding of E. coli O157:H7 by infected dairy cattle appears to be transient and persistence of E. coli O157:H7 was not demonstrated from the farm environment sites tested.     

大肠埃希菌O157:H7的金增强胶体金免疫渗滤检测法

目的 建立基于盐酸羟胺晶种法的金增强的免疫渗滤法,快速、灵敏检测大肠埃希菌(Escherichia coli)0157:H7.方法 以柠檬酸钠还原法制备粒径约为18nm的免疫胶体金;采用羟胺晶种法对胶体金粒子进行生长放大,考察金生长液中盐酸羟胺与氯金酸的物质量的比对胶体金粒子生长放大的影响;借助透射电镜观察胶体金粒子在...     

出血性大肠杆菌O157∶H7的脉冲场凝胶电泳分型方法研究

目的应用脉冲场凝胶电泳技术(PFGE)对肠出血性大肠杆菌O157:H7进行基因分型,了解不同来源分离株的相关性。方法选择在广西食品污染物检测地区食品样本中分离到的7株出血性大肠杆菌O157:H7,采用琼脂糖包埋胶块法制备基因组DNA,以XbaⅠ限制性内切酶进行酶切,选定脉冲电泳参数进行脉冲场凝胶电泳,照相记录结果;Nei的公式计算相似性和遗传距离,并用PHYLIP程序非加权成对算术平均法聚类分析。结果PFGE可生成14~18条清晰条带的分离株DNA指纹图谱,由聚类分析树状图可将7株分离株分为6个基因群。结论广西不同地区食品分离O157:H7血清型菌株存在多个分子亚型,提示不是一个克隆株,研究发现脉冲场凝胶电泳分型技术重复性高,在分子流行病学研究中有较大的应用前景。